Astrocytes of the vertebrate CNS have been proposed to perform a wide variety of different functions including acting as a substrate for axon growth during development, contributing to the induction of the blood brain barrier and forming glial scars following injury to the adult CNS. Our recent studies suggest that astrocytes are a heterogeneous class of cells. In the proposed studies we will examine astrocyte diversity in the rat spinal cord and test the hypothesis that distinct types of astrocytes perform distinct functions in the developing, adult and injured spinal cord. Three different approaches will be used to define astrocyte diversity in the developing spinal cord: single cell cloning, retrovirus mediated gene transfer and the generation of cell type specific monoclonal antibodies. Initial studies suggest that cultures of neonatal rat spinal cord contains five morphologically distinct types of astrocytes. We will determine the time of origin, and the factors, that regulate proliferation and differentiation of each of these different cell types. Retrovirus mediated gene transfer will be used to determine the cell lineage relationships of the different types of astrocytes, both in cultures containing all classes of spinal cord cells and in the intact developing spinal cord. Recently isolated, as well as novel cell-type specific monoclonal antibodies will be used to distinguish biochemical differences between these cell types and to localize them in the intact spinal cord. The functional potential of the different types of astrocytes will be examined by analyzing their ability to support neurite outgrowth from developing neurons, suppress glial scar formation and reestablish the blood brain barrier after injury in the adult. The extent of neurite outgrowth and its molecular basis on the different populations of astrocytes will be established using purified cell populations in vitro. Transplantation of nitrocellulose implants coated with the different populations of astrocytes in to the adult CNS will be used to assess the ability of each type of cell to suppress glial scar formation and re-establish the blood brain barrier after injury. These studies will provide new, and important information on the extent and functional significance of astrocyte diversity in the spinal cord. Since the response of astrocytes to CNS injury is important for functional recovery it is critical to define specific astrocytic cell types in the CNS and to describe their individual functions following a CNS lesion. Such information is essential if we are to understand and subsequently manipulate the complex astrocytic response to CNS injury.